This is a continuation of our studies of the electrical behavior of sympathetic preganglionic neurons (SPN's) in vitro and of transmitters activating or modulating the activity of these neurons. Experiments will be performed on SPN's contained in thin (300-400 Mum) sections of thoracic spinal cord removed from neonatal (10-15 days old) rats and from age-matched spontaneously hypertensive rats (SHR); intracellular current or voltage recording techniques will be used. Fluorescence labeling of SPN's in thin slices will be accomplished by placing for 1-2 hours a drop of 3% lucifer yellow CH solution on ventral rootlets. Spinal cord slices will be superfused with a physiological solution and viewed under a fluorescent microscope. Glass microelectrodes will be inserted under visual guidance into fluorescent SPN's. The proposed research has five major objectives. First, single electrode voltage clamp will be employed to voltage clamp SPN's with the aim to characterize the membrane currents that can be recorded from soma of SPN's. Second, synaptic potentials will be evoked by bipolar stimulating electrode placed 100-200 Mum away from the impaled SPN, and their characteristics will be analyzed under current or voltage clamp conditions. The data obtained will serve as a basis for identification of the transmitters responsible for synaptic potentials. Third, pharmacological antagonists to amino acids, biogenic amines and peptides will be employed to identify which transmitters may be involved in the generation of evoked synaptic potentials; furthermore, attempts will be made to mimic the synaptic potentials by application of suspected transmitters. Fourth, effects on SPN's and on evoked SPN transmission of centrally acting antihypertensive drugs such as clonidine and methyldopa, will be evaluated for their possible spinal site of action. Fifth, electrical membrane properties and chemosensitivity of the SPN's of spinal cord slices obtained from SHR will be investigated and compared with those of SPN's obtained from normal rats. The significance of this research is threefold: (1) this study will lead, for the first time, to characterization of the transmitters and modulators acting on SPN's; this will give insight into circuitry of supra-spinal control of the sympathetic nervous system; (2) comparison of electrical membrane properties of SPN's of normal rats and SHR and of their chemosensitivity will help our understanding of hypertension; and (3) identification of the site and mechanism of action of clonidine and methyldopa will establish the spinal cord as a potential site of action of antihypertensive drugs.